In 1992, Semenza et al. found a protein capable of specifically binding to hypoxia responsible element of erythropoietin gene, and this protein is named as Hypoxia inducible factor-1 (HIF-1). HIF-1 widely exists in mammalian cells, and mainly regulates angiogenesis, cellular survival and death, as well as pH. HIF-1 is a heterodimer consisting of two subunits (HIF-α and HIF-β) and both of these two subunits are members of alkaline helix-loop-helix transcription factor superfamily, and have PER-AHR/ARNT-SIM (PAS) domain. HIF-α has 3 subunits, which are separately HIF-1α, HIF-2α and HIF-3α, in which HIF-1α and HIF-2α have a high sequence identity, recognize the same DNA binding domain, but have different biological effects.
Under normoxia condition, proline residues at sites 402 and 564 in oxygen-dependent degradation domain (ODDD) of HIF-1α are hydroxylated by prolyl hydroxylase (PHD), and collect ubiquitin proteins such as elongin C, elongin B to form ubiquitin-linking protease complexes and to be degraded via ubiquitin-dependent pathway when they bind to tumor suppressor von Hippel-Lindau gene product (pVHL). Under anoxic conditions, deactivation of PHD results in that proline residues cannot be hydroxylated, HIF-1α cannot bind to pVHL, so that its degradation is hindered.
Activation of HIF-1α results in upregulation of downstream target genes, and this has been confirmed in more than 70 HIF-1α target genes, which widely influence many physiological phenomena such as carcinogenesis, vasculogenesis, cell survival, glucose metabolism, etc. Among the downstream target genes, vascular endothelial growth factor (VEGF) and erythropoietin (EPO) are key growth factors for tumor angiogenesis, glucose transporter (Glut-1) can reduce glucose level, carboanhydrase IX (CAIX) can regulate pH change.
In 2001, Bruick found a protein capable of hydroxylating HIF-α, and then Ivan et al found genes encoding proline hydroxylase in human genes, i.e., EGLN-1, EGLN-2 and EGLN-3, which encode proteins PHD2, PHD1 and PHD3. In 2002, Oehme found the 4th PHD, i.e., PHD4, which could hydroxylate HIF-α when it was expressed at a high level.
According to inhibition mechanism of proline hydroxylase, they are roughly classified into two groups, i.e., iron chelating agents or iron competitive agents and 2-ketoglutaric acid analogues. Iron chelating agents or iron competitive agents can bind to Fe2+ binding site of ODDD, thereby preventing HIF-α from binding to proline hydroxylase (e.g., prolyl-4-hydroxyases; EC 1.14.11.2), terminating hydroxylation reaction, avoiding degradation of HIF-α, and achieving stabilization of HIF-α. 2-Ketoglutaric acid analogues are compounds capable of inhibiting proline hydroxylase via competition with endogenous 2-oxoglutarate (2-OG), and currently, these inhibitors for proline hydroxylase are research focus.
At present, hot research relates to synthesizing peroral small molecular inhibitors for proline hydroxylase, to stabilize HIF-1a, and thus upregulating its downstream target genes, including but not being limited to erythropoietin (EPO), hemeoxygenase-1, adiponectin inducible nitric oxide synthase), etc., thereby achieving purpose of treating diseases such as anemia, acute ischemic reperfusion injury, etc.